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Abstract: FR-PO865

Inhibition of Cyclophilin A and Large T Antigen Interaction Reduces Polyomavirus BK Replication

Session Information

  • Transplantation: Basic
    October 26, 2018 | Location: Exhibit Hall, San Diego Convention Center
    Abstract Time: 10:00 AM - 12:00 PM

Category: Transplantation

  • 1801 Transplantation: Basic

Author

  • Tian, Ya-chung, Chang-Gung Memorial Hospital, Taipei County, Taiwan

Group or Team Name

  • Linkou Kidney Research Center
Background

Specific treatment for polyomavirus BK (BKV) infection in kidney transplant recipients remains unsolved. We previously demonstrated that cyclophilin A (CypA) was crucial for BKV replication. The aim of this study was to assess the impact of interaction of CypA and BKV large T antigen (TAg) on BKV replication.

Methods

HRPTEC or HK-2 were used in this study.

Results

The immunoprecipitation (IP) revealed an interaction between TAg and CypA. Similarly, the ELISA assays using purified TAg and CypA confirmed a binding between CypA and TAg. Atomic force microscope (AFM) revealed a binding between TAg and CypA. This binding was reduced if the TAg-coated probe was pretreated with anti-TAg blocking antibody. Immunocytochemistry (IHC) also verified nuclear colocolization of CypA and TAg expression. Both N-terminus (DN-CypA) and C-terminus (DC-CypA) failed to colocalize with nuclear TAg expression. The IP assay confirmed a failure of DN-CypA binding to TAg. Overexpression of DN-CypA in the CypA-knockdown cells reduced BKV replication, confirming that the N-terminus of CypA binding to TAg is crucial for viral replication. Overexpression of CypA but not DN-CypA enhanced TAg-induced promoter activity of BKV. Overexpression of CypA in the NFATc3-knockdown cells still enhanced viral replication, indicating a NFATc3-independent pathway. Addition of Debio-025 caused a reduction in BKV replication. IHC showed that Debio-025 caused a reduction of nuclear CypA expression. Immunoblotting assay confirmed a dramatic reduction of CypA in the nuclear fraction by addition of Debio-025. Mutant CypA proteins at six different functional residues in the enzyme pocket of CypA (possible functional sites of Debio 025) did not affect TAg and CypA interaction detected by the IP assay and the ELISA assay. In vivo, following BKV injection, BKV copies in the kidneys were reduced in the CypA-knockout mice or in the Debio-025-treated mice, when compared with those in the wild-type mice or non-treatment controls.

Conclusion

Nuclear translocation of CypA and interaction with nuclear TAg, both of which depends on N-terminus of CypA is essential for BKV replication. Blockade of the CypA-dependent NFAT-independent pathway by Debio-025 suppressed BKV replication. As no anti-BKV treatment is satisfactory so far, alisporivir that has no immunosuppressive effect can be potentially applied for anti-BKV treatment.

Funding

  • Government Support - Non-U.S.